Distinct effects of flow intermittency on the benthic microbial diversity and their denitrification on different substrates

Global climate change has significantly increased the duration of droughts in intermittent rivers, impacting benthic microbial-mediated biogeochemical processes. However, the response mechanisms of biofilms on different substrate types to alternating dry and wet conditions and their related ecosystem functions remain poorly understood. This study uses high-throughput sequencing and enzyme assays to investigate the impact of gradient drought stress on microbial diversity and functional changes of biofilm communities inhabiting on gravel, cobblestone, and sediment. Results showed that the duration of drought significantly affects microbial diversity, with algal and bacterial α-diversity declining under extended drought across gravel, cobblestone, and sediment substrates. At the same time, fungal diversity was less affected, likely due to their distinct ecological niches and reproductive strategies. β-diversity analysis revealed significant changes in community heterogeneity, with algae and bacteria showing increased Bray-Curtis dissimilarities, indicating distinct adaptation strategies that may affect ecosystem functioning. Fungal communities, however, were less impacted by drought-induced heterogeneity changes. Network analysis showed that drought altered microbial network connectivity, with algal networks displaying decreased path distances, while bacterial networks remained stable, suggesting greater resilience to drought stress. Functional enzyme assays revealed significantly reduced denitrification rates across all substrates post-drought, with distinct denitrifying enzyme activity responses depending on substrate type. Partial least squares path modeling revealed that algal biodiversity were closely linked to the maintaining of enzyme activities, particularly denitrification rates of biofilms on cobblestone and gravel. These findings indicated the critical role of substrate types in shaping microbial responses to drought stress, with distinct microbial groups and diversity indices playing key roles in maintaining ecosystem functions. This study highlights the importance of understanding the interactions between microbial community dynamics and ecosystem functions under varying environmental stressors in river ecosystems.